Seasonal reproductive and metabolic changes in male Pacific white-sided dolphins (Aethalodelphis obliquidens)

Seasonal reproductive dynamics were investigated in two adult male Pacific white-sided dolphins (Aethalodelphis obliquidens, formerly Lagenorhynchus obliquidens) maintained at the Enoshima Aquarium over a two-year period (December 2018-November 2020). Serum testosterone (T), testicular cross-sectional area (TS), and pixel intensity (PI), an index of testicular echogenicity, were measured along with rectal body temperature at two depths (shallow, 15 cm; deep, 30 cm). Untargeted serum metabolomic profiling was performed using gas chromatography-mass spectrometry (GC-MS). In both individuals, T showed a clear seasonal pattern, with elevations lasting approximately one month during spring and summer, followed by prolonged increases in TS (approximately three months) and PI (three-four months). This temporal sequence was consistent across both years of observation. The T, TS, PI, and photoperiod were strongly and positively correlated with one another (Spearman's rs > 0.74, P < 0.01). Shallow-body temperature varied seasonally with water temperature, whereas deep-body temperature remained relatively stable. GC-MS detected 131 metabolites, and partial least squares discriminant analysis revealed a separation between high-T (> 10 ng/ml) and low-T (< 1 ng/ml) states. The four metabolites with the highest variable importance in projection scores were myoinositol, palmitoleic acid, L-tyrosine, and L-phenylalanine. Myoinositol and palmitoleic acid were positively correlated with the reproductive indices, whereas L-tyrosine and L-phenylalanine were negatively correlated, suggesting coordinated shifts in inositol, lipid, and amino acid metabolism during reproductive activation. To the best of our knowledge, this is the first study to integrate endocrine, ultrasonographic, thermal, and metabolomic approaches to characterize male reproductive seasonality in this species.